Chronic dysregulation of physiological systems manifests systemically as allostatic load (AL) and in abnormal stress reactivity profiles, which are features of psychopathological conditions that increase future disease risk. But the biological basis underlying inter-individual differences in stress regulation and reactivity remains unknown. Even among populations of healthy individuals exposed to standardized laboratory challenges, such as the Trier social stress test (TSST), there are substantial differences in the magnitude of responses in multiple physiological systems, including but not limited to the hypothalamic-pituitary-adrenal (HPA) axis, the autonomic nervous system (ANS) and cardiovascular system, metabolic changes, and immune and pro-inflammatory systems. One common factor to all stress systems is their dependence on energy supply, which fuels every aspect of the stress response including molecular, cellular, systemic and cognitive/psychological functioning. At the cellular level, energy is provided by mitochondria, unique organelles that populate the cell cytoplasm and contain their own genome, the mitochondrial DNA (mtDNA), that is essential to mitochondrial health. MtDNA defects cause dysregulation of multiple aspects of mitochondrial structure and function, known as mitochondrial allostatic load (MAL). Three main lines of evidence suggest that MAL contributes to AL and regulates stress responses in humans: i) we recently discovered that the mtDNA is released following psychological stress in humans (PNEC 2019), ii) experimentally-induced MAL in animals caused specific alterations in the multisystem physiological responses to psychological stress (PNAS 2015), and iii) mitochondria are the source of stress hormones, including cortisol that is synthesized in mitochondria within the adrenal glands (Nat Genetics 2012). Together, this evidence suggests that MAL may alter both baseline AL and stress reactivity profiles, potentially providing new insight into the source of interindividual differences in stress regulation and health in general. In this project, we perform the first comprehensive assessment of MAL, systemic AL, multisystem stress reactivity to a laboratory challenge (TSST) in three groups of individuals who have rare genetic mtDNA defects that selectively causes different forms of MAL and in a healthy control group. Multisystem stress biomarker profiling under fasting baseline and stress reactive conditions will provide a comprehensive test of pathways linking MAL to stress physiology in humans. The resulting high-dimensionality data will be treated using integrative data analytic approaches and classifying algorithms, including cross-validated machine learning models, to identify resting and stress-reactive biomarker signatures responsive to MAL. In parallel, assessments of executive function and key domains of psychosocial functioning including mood, stress, anxiety, depressive symptoms, and well-being will contribute to provide a comprehensive picture of novel mitochondrial psychobiological pathways.